The major objective of this project is the elucidation of the mechanism and the regulation of the interaction of mRNA with the ribosomes. In previous years we have focused on the properties and functions of ribosomal protein SI from E. coli and an analogous protein from Caulobacter crescentus. SI was shown to be indispensible for ribosomal binding of natural mRNA. This function of SI was shown to be related to its RNA helix-destabilizing (unwindling) properties. In the last year emphasis was shifted toward analogous problems in eukaryotes. A ribosome-associated unwinding protein was isolated from embryos of Artemia salina and purified to homogeneity in a six-step procedure. The protein, UP-40 (MW 40,000) is in many respects analogous to DNA helix-unwinding proteins, as is SI. UP-40 inhibits protein synthesis in cell-free systems; inhibition is relieved by an excess of mRNA. These properties are again analogous to the properties of SI. Undeveloped A. salina embryos contain metabolically inactive monosomes and mRNA-protein particles. Upon resumption of development (aeration in a salt solution) protein synthesis is reinitiated. We find that the amount of UP-40 associated with the ribosomal fraction decreases drastically upon development. An antibody against UP-40 is being raised and should, hopefully, answer questions related to the precise location of UP-40 in subcellular fractions (ribosomes, mRNA-protein particles, etc.): it should also help in investigations on the in vivo function and fate of UP-40 (degradation, modification, etc.) during development. The function of this protein, and other RNA binding proteins associated with ribosomes and/or with mRNA protein particles in dormant eggs and their changes during development are the major problems for the coming year.